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Recombinant firefly luciferases and their applications for the detection of microorganisms Satoshi Fukuda*; S Murakami; H Tatsumi R & D Division, Kikkoman Corp., 399 Noda, Noda City, Chiba 278-0037, Japan      *(sfukuda@mail.kikkoman.co.jp) Firefly luciferase based assays have been widely used to monitor sanitation conditions by detecting ATP. Kikkoman succeeded in the cloning and expression in Escherichia coli of cDNA clones of luciferases from two kinds of Japanese firefly (Luciola lateralis and L. cruciata). These cDNA clones made it possible for us to improve the properties of luciferases by genetic engineering. We constructed thermostable mutants that contributed to the improved stability of products. Moreover, we have obtained a second generation of recombinant luciferases for practical properties. One is a detergent resistant mutant, which enabled us to use detergent at a higher concentration than before. Using this mutant in combination with ATP eliminating enzymes and a luciferin derivative, respectively, we developed kits for microbial biomass assay and coliform testing. Another one is biotinylated luciferase, which is a fusion protein between a thermostable luciferase and a biotin acceptor peptide. We also developed a rapid Staphylococcus aureus screening kit using the biotinylated luciferase and specific antibodies. [Talk: fukuda.satosh.86903]

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